We describe the synthesis and results of biological evaluation of newly designed 2,4,6-trisubstituted symmetrical 1,3,5-triazine (TAZ) derivatives. Among the tested trisubstituted TAZ derivatives, some C_S-symmetrical alkoxy-amino-substituted TAZ derivatives, including 7ggp and 6dpp, showed significant antiviral activity against herpes simplex virus type 1 (HSV-1). The compound with the highest level of antiviral activity was C₃-symmetrical trialkoxy-TAZ derivative 4bbb, which showed a considerably high selectivity index (IC₅₀/EC₅₀=256.6). The structure–activity relationships for anti-HSV-1 activity of the tested 2,4,6-trisubstituted TAZ derivatives are also described.

Supramolecular interaction of two-fold (C₂) or three-fold (C₃) symmetrical geometry macromolecules with many bioactive compounds is one of the common features of many important biological processes,^1–3) and small molecules having C₃-, C_S-, or C₂-symmetrical geometry often appear in various biologically active compounds contrasted on a symmetrical template.^4–6) We have therefore expected that such small symmetrical molecules would be promising new candidates or leads in the search for biologically active compounds that interfere with the sugar recognition process in a controlled biological response. From this aspect of molecular symmetry, we have already reported a few examples of such types of new symmetrical molecules for the purpose of finding new biological active compounds.^7–9)

We have recently reported some molecular modifications of 2,4,6-trichloro-1,3,5-triazine (TCTAZ) to C₃-, or C_S-symmetrical trisubstituted TAZ molecules and the results of biological evaluation of synthesized symmetrical 2,4,6-trisubstituted TAZ derivatives.⁷⁾ Among previously targeted TAZ derivatives, some alkoxy-amino-substituted derivatives showed significant anti-herpes simplex virus type 1 (HSV-1) activities. The previous analysis of anti-HSV-1 active molecular features indicated that a C_S-symmetrical TAZ derivative with two alkoxy groups and one amine moiety seemed to be a required structure for preferred anti-HSV-1 activity with a good selectivity index.

For an extension of our study, we examined further modifications of this symmetrical class of compounds in order to investigate the structure–activity relationship (SAR) of alkoxy-amino-substituted TAZ molecules as well as their biological evaluation. In this paper, we report the results of preparation of newly targeted C_S-symmetrical TAZ derivatives together with the results of biological evaluation of obtained symmetrical 2,4,6-trisubstituted TAZ derivatives.

In our previous article,⁷⁾ we reported that synthesis of target alkoxy-amino-trisubstituted TAZ derivatives is easily achieved by a procedure via alkoxy-substituted chloro-TAZ intermediates starting from TCTAZ (1). This procedure consists of two-stage nucleophilic substitutions of compound 1 with alcohols (Step 1) and then amines (Step 2) in one pot. The predominant formation of a monoalkoxy-diamino-TAZ derivative (6) or dialkoxy-monoamino-TAZ derivative (7) in an employed reaction depended on the stoichiometry of TCTAZ and alcohol in the first reaction stage (Step 1). As an amine nucleophile for introduction on a TAZ template in this study, we employed only 4-piperidinemethanol (pH) because many molecules having this amine substituent designed previously showed considerable antiviral activities.

The overall reaction stage for the preparation of target molecules (6 and 7) is shown in Chart 1 together with obtained by-products. The results of the reactions of TCTAZ (1) with various alcohols (ROH: a–gH) and 4-piperidinemethanol (pH=XH) are summarized in Table 1.

Table 1. Synthesis of Alkoxy-amino-TAZ Derivatives (6 and 7) from Reactions of TCTAZ (1) with Various Alcohols (ROH) and 4-Piperidinemethanol (pH)^a)

Entry	ROH	Methodb)	Ratio of 1 : ROH : (Additive 1) : pH : (Additive 2)	Conditions in Step 1)	Conditions in Step 2)	Products (yield %)c,d)
1	aH	A	1 : 1.2 : (1.2) : 4 : (4)	0°C 1 h	rt 14 h	6app (61), 7aap (trace)
2	aH	A	1 : 2.4 : (2.4) : 2 : (2)	0°C 1 h	rt 14 h	6app (79), 7aap (3), 8ap (8)
3	aH	A	1 : 2 : (8) : 2	0°C 15 min to rt 16 h, N2, THFe)	70°C 1 h, N2, THFe)	6app (13), 5pp (1), 8ap (62)
4	aH	A	1 : 25.5 : (3) : 2 : (2)	rt 1 h, N2	rt 1.5 h to reflux 39 h, N2, acetonee)	6app (21), 8ap (8)
5	aH	A	1 : 10 : (5) : 2 : (2)	rt 0.5 h to 60°C 2.5 h, N2	rt 18 h, N2	6app (8), 7aap (60)
6	bH	A	1 : 10 : (2) : 2 : (2)	rt 0.5 h to reflux 4 h, N2	rt 21 h	6bpp (84)
7	bH	A	1 : 10 : (2) : 2 : (2)	rt 1 h to reflux 19 h, N2	rt 21 h	6bpp (73), 7bbp (26)
8	bH	B	1 : 4.8 : (3)	rt 1 h to reflux 17 h, CHCl3e)	—	2b (28)
9	bH	B	1 : 6 : (3)	rt 7 d, N2, THFe)	—	3bb (2), 4bbb (9)
10	cH	A	1 : 10 : (5) : 2 : (2)	rt 30 min, N2	rt 23 h, N2	6cpp (72)
11	cH	A	1 : 10 : (2.5) : 2 : (2)	rt 1 h to reflux 22 h, N2	rt 38 h	6cpp (12), 7ccp (10)
12	cH	C	1 : 2 : (2) : 1.6	rt 30 min to reflux 2.5 h, N2	rt 30 min	7ccp (73)
13	dH	A	1 : 2.2 : (2.2) : 2 : (2)	rt 1 h to reflux 16 h, N2	rt 24 h	6dpp (62), 7ddp (6)
14	dH	A	1 : 10 : (2) : 2 : (2)	rt 2.5 h to reflux 17 h, N2	rt 21 h	6dpp (67), 7ddp (25)
15	dH	C	1 : 2 : (2) : 1.6	rt 30 min to reflux 2 h, N2	rt 30 min	7ddp (67), 5pp (detected)
16	eH	A	1 : 10 : (5) : 2 : (2)	rt 1 h to reflux 3 h, N2	rt 19 h	6epp (80)
17	eH	A	1 : 10 : (2) : 2 : (2)	rt 1 h to reflux 1 d, N2	rt 20 h	6epp (22), 7eep (54)
18	fH	A	1 : 2.2 : (2.2) : 2 : (2)	rt 1 h to reflux 19 h, N2	rt 23 h	6fpp (54), 7ffp (38)
19	fH	A	1 : 10 : (2) : 2 : (2)	rt 1 h to reflux 20 h, N2	rt 4 h	6fpp (31), 7ffp (52)
20	gH	A	1 : 2 : (2) : 2 : (2)	rt 5.5 h	rt 2 h	6gpp (4), 7ggp (47)
21	gH	A	1 : 2.6 : (2) : 2 : (2)	rt 2 h	rt 4 d, acetonee)	6gpp (16), 7ggp (52)

a) DIPEA stands for N,N-diisopropylethylamine. Dry solvents were used in all reactions except for CHCl₃ in the reaction of Entry 8. b) The additives and solvents used are as follows: Method A (Additive 1=collidine, Solvent 1=acetone, Additive 2=DIPEA, Solvent 2=CH₃CN), Method B (Additive 1=NaHCO₃, Solvent 1=CHCl₃ or THF), and Method C (Additive 1=n-BuLi, Solvent 1=THF, no Additive 2, Solvent 2=CH₃CN). c) Yield obtained from TCTAZ (1). d) The by-products 6wpp (16%) and 7wap (12%) [Entry 1], 6wpp (14%) [Entry 2], 6wpp (25%) [Entry 5], 7wbp (5%) [Entry 6], 7wep (2%) [Entry 17], and 7wfp (5%) [Entry 18] were also isolated. These compounds apparently suffered from hydrolysis of intermediates by moisture in the commercial alcohols used in each entry. In Entry 3, the by-product 5rp (structure shown below) derived from the reaction with DIPEA was also isolated in 14% yield. e) Solvent used in this step.

Most of the reactions were performed first by our reported procedure (Method A), in which we used collidine and DIPEA as bases to trap HCl generated in both reaction stages (Steps 1 and 2). As shown in Table 1, target monoalkoxy-diamino-TAZ derivatives (6app-fpp) were obtained in good to excellent yields from the various reactions under conditions using Method A (Entries 1, 2, 6, 11, 13, 14, 16, and 18) with an excess amount of alcohols ROH (a–fH).

Chart 1. Synthesis of 2,4,6-Trisubstituted TAZ Derivatives

Among the compounds isolated from reactions of applied entries (Entries 1 and 2), a few products from the reactions of TCTAZ and water were isolated (see compounds 6wpp and 7wap). When the reactions were conducted in dry tetrahydrofuran (THF) or dry acetone in both Steps 1 and 2 under a N₂ atmosphere (Entries 3 and 4) using Method A, there was a tendency for decrease in yields of the target compounds (6app and 7aap).

To improve the yield of target dialkoxy-monoamino-TAZ derivatives (7), we tried the procedure of Method A with various conditions. In the reactions with a primary alcohol aH, improvement in the yield of product 7aap was achieved by the reaction using a large amount of alcohol and a base as an additive at a slightly elevated temperature (60°C) in Step 1 and using a long reaction period (18 h) in Step 2 (Entry 5). However, reactions under similar conditions with a branched-chain secondary alcohol such as bH or cH (Entries 6 and 7, or Entries 10 and 11) gave target dialkoxy-substituted TAZ derivatives (7bbp or 7ccp) in low yields (26 and 10% in Entries 7 and 11, respectively) or resulted in no isolation of such target compounds (Entries 6 and 10). On the other hand, in reactions using other cyclic secondary alcohols (d–fH) having less steric hindrance than branched-chain secondary alcohols (Entries 14, 17, and 19), all target TAZ derivatives (7ddp, 7eep, and 7ffp) were obtained in 25, 54, and 52% yields, respectively. The tendency observed in these reactions was that the use of cyclic secondary alcohols results in better yields of target TAZ derivatives (7) than the yields when branched-chain secondary alcohols are used.

To improve the yield of the target TAZ derivative 7bbp, we further applied a similar method to the reported procedure using NaHCO₃^10,11) (Method B) to obtain an intermediate 3bb; however, a few of our trials were unsuccessful and resulted in the isolation of monoalkoxy-dichloro-TAZ intermediate 2b¹²⁾ (Entry 8)¹⁰⁾ or trialkoxy TAZ (4bbb)¹³⁾ (Entry 9)¹¹⁾ as a main product in low yields.

With the aim of improvement of the yield of the target dialkoxy TAZ derivatives (7), we also attempted reactions with secondary alcohols (cH and dH) (Entries 12 and 15) and found that the use of n-BuLi¹⁴⁾ (Method C) is effective for the formation of the desired dialkoxy-TAZ derivatives (7ccp and 7ddp). The generated alkoxide anion (RO⁻) probably works as a better nucleophile to give the target TAZ derivatives (7) in high yields. For the preparation of dialkoxy-monoamino-TAZ derivatives (7), this method has a considerable advantage over the others (Methods A and B) in terms of reaction yield.¹⁵⁾

In the case of a phenolic alcohol (sesamol: gH), the target derivative (7ggp) was smoothly obtained by reactions of TCTAZ (1) with gH (Entries 20 and 21) using Method A, together with the formation of a small amount of monoaryloxy-TAZ derivative (6gpp).

From these reactions of TCTAZ with various alcohols (a–gH) and an amine (pH) described above, we could achieve conventional preparation for targeted symmetrical TAZ derivatives (6 and 7). In some runs, we also isolated other trisubstituted TAZ derivatives including 4bbb, 6wpp, 7wap, 7wbp, 7wep, 7wfp, and 8ap formed as by-products (see Table 1).

All structures of the synthesized compounds were easily confirmed by spectroscopic and analytical data. The geometries of C₃- or C_S-symmetrical structures of target TAZ derivatives described in this article were also confirmed by ¹³C-NMR spectroscopic data.

With respect to a three-dimensional interaction of a bioactive molecule for its binding site, it is expected that van der Waals interactions or formation of hydrogen bonds between substituents in the bioactive molecule and a host macromolecule play an important role for biological activity. The results obtained in our previous experiments⁷⁾ seemed to indicate that a C_S-symmetrical TAZ structure with an amine and/or two alkoxy groups on a TAZ template is required for anti-HSV-1 active interaction.

The structures of targeted C_S-symmetrical 2,4,6-trisubstituted TAZ derivatives in this study obtained from TCTAZ and results of biological evaluations [anti-HSV-1 activities (EC₅₀) by plaque reduction assays¹⁶⁾ and their cytotoxicities against Vero cells (IC₅₀)] are summarized in Table 2 together with the data of aciclovir.¹⁷⁾ Calculated log P values¹⁸⁾ for the compounds are also shown in Table 2. There were few distinct correlations between log P values and EC₅₀ values or between log P values and IC₅₀ values among the compounds listed in Table 2.

Table 2. Antiviral Activity (EC₅₀), Cytotoxicity (IC₅₀) against HSV-1, and Calculated Log P

Compound		EC50 (μM)	IC50 (μM)	Log Pa)	Compound		EC50 (μM)	IC50 (μM)	Log Pa)
	6app	>100	>200	1.93		6fpp	>25	81.2	4.05
	7aap	>100	>200	1.43		7ffp	>12.5	27	5.67
	6bpp	>100	>200	2.74		6gppb)	>6.3	42.2	3.52
	7bbp	>100	>200	3.05		7ggp	32.2	303.6	4.62
	6cpp	>100	116.8	3.71		4bbb	1.87	479.8	3.36
	6dpp	67.2	295.4	3.21		6wpp	>100	>200	1.82
	7ddp	>100	28	4.00		7wap	>100	>200	1.32
	6epp	124.3	224.9	3.63		7wfp	>100	>200	3.44
	7eep	>100	85.1	4.83		8ap	>100	>200	1.89
					Aciclovirc)		1.1	>444	−0.76

a) Log P was calculated by using ChemBioDraw 13.0.2.3021. b) Data were taken from ref. 7. c) Data were taken from ref. 17.

C_S-symmetrical derivatives (6app and 6bpp) having a relatively small aliphatic alkoxy group showed no significant anti-HSV-1 activity (EC₅₀=>100 µM) or cytotoxic activity (IC₅₀=>200 µM). Introduction of two same alkoxy groups in the above TAZ template (7aap and 7bbp) also resulted in no significant biological activities, indicating no enhancement effect of this modification. However, modifications of TAZ derivatives having a more bulky alkoxy group than the isopropoxy group in the TAZ template (such as 6dpp, 6epp, and 6fpp) to TAZ derivatives having two bulky alkoxy groups in the template (such as 7ddp, 7eep and 7ffp) resulted in C_S-symmetrical compounds with greater cytotoxicity as shown in Table 2. In contrast, a similar modification of compound 6gpp by introduction of an aryloxy group resulted in reverse cytotoxic activity affording compound 7ggp with lower cytotoxicity.

Anti-HSV-1 activities of some other 2,4,6-trisubstituted TAZ derivatives (4bbb, 6wpp, 7wap, 7wfp, and 8ap) obtained through our synthetic trials were also evaluated and the results are shown in Table 2. It is noteworthy that the C₃-symmetrical TAZ derivative 4bbb showed surprisingly high anti-HSV activity (EC₅₀=1.87 µM) and low cytotoxic activity (IC₅₀=479.8 µM), thus resulting in a better selectivity index (IC₅₀/EC₅₀=256.6). This unexpected result provides useful information for modification of this symmetrical class of compounds.

Triamino-substituted C₃-type derivatives reported in our previous paper⁷⁾ showed no anti-HSV activity at a concentration less than 100 µM. On the basis of previous data, we considered the dialkoxy-amino-substituted TAZ structure to be a probable core structural feature for an anti-HSV-1 active molecule for this series. In terms of high anti-HSV-1 activity with a good selectivity index, new trialkoxy-substituted TAZ derivatives such as 4bbb appear to have a great potential as leads in the search for antiviral compounds. In order to verify this potential information, we have already begun to perform further various structural modifications of compound 4bbb and biological evaluation of the TAZ derivatives obtained as a new project. We are also investing the sugar recognition properties of highly antiviral active tri-substituted TAZ derivatives by using isothemal titration calorimetry. The results of modification and SAR studies of the trialkoxy-substituted TAZ derivatives will be described in the following paper.

Melting points were determined using a micro melting point apparatus (Yanaco MP-S3) without correction. IR spectra were measured by a Shimadzu FTIR-8100 IR spectrophotometer. Low- and high-resolution mass spectra (LR-MS and HR-MS) were obtained by a JEOL JMS HX-110 double-focusing model equipped with an FAB ion source interfaced with a JEOL JMA-DA 7000 data system. ^lH- and ¹³C-NMR spectra were obtained by JEOL JNM A-500. Chemical shifts were expressed in δ ppm downfield from an internal TMS signal for ^lH-NMR and the carbon signal of the corresponding solvent [CDCl₃ (77.00 ppm), DMSO-d₆ (39.50 ppm), and THF-d₈ (68.60 ppm)] for ¹³C-NMR. The abbreviations qu=quintet, dm=double multiplets and tm=triple multiplets are used for the multiplicity of ^lH-NMR data, respectively. The signal assignments were confirmed by two-dimensional (2D)-NMR analyses: ^lH–^lH 2D correlation spectroscopy (COSY), ^lH–^l3C heteronuclear multiple-quantum coherence (HMQC), ^lH–^l3C heteronuclear multiple-bond connectivity (HMBC). Microanalyses were performed with a Yanaco MT-6 CHN corder. Routine monitoring of reactions was carried out using precoated Kieselgel 60F₂₅₄ plates (E. Merck). Centrifugal or flash column chromatography was performed on silica gel (Able-Biott or Kanto 60N) with a UV detector. Commercially available starting materials were used without further purification, and dry solvents were used in all reactions except for Entry 8.

To a solution of 2,4,6-trichlorotriazine (TAZ: cyanuric chloride) (1, 922 mg, 5.0 mmol) and collidine (727 mg, 6.0 mmol) in dry acetone (10 mL) was added 2-methoxyethanol (aH, 457 mg, 6.0 mmol) at 0°C. After stirring for 1 h at 0°C, the resulting colorless precipitated collidine·HCl was removed by filtration and then the solvent was evaporated to afford a yellow oily residue. (Step 2) This material was dissolved in dry CH₃CN (15 mL), and 4-piperidinemethanol (dH, 2.30 g 20.0 mmol) and N,N-diisopropylethylamine (DIPEA, 2.58 g, 20.0 mmol) were added, and the resulting mixture was stirred for 14 h at room temperature. After evaporation of the solvent, the residue was separated by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=93 : 6.5 : 0.5→85 : 14.5 : 0.5) to give 6app (1.17 g, 61% yield) as a yellow solid. Compounds 6wpp [4,6-bis[4-(hydroxymethyl)piperidin-1-yl]-1,3,5-triazin-2(1H)-one ] (253 mg, 16% yield) as a pale yellow solid and 7wap [4-[4-(hydroxymethyl)piperidin-1-yl]-6-(2-methoxyethoxy)-1,3,5-triazin-2(1H)-one] (173 mg, 12% yield) as a colorless opaque solid were also isolated in reaction products. Compound 7aap was also detected by TLC. Recrystallization from EtCN or EtOH gave an analytically pure product 6app or 6wpp.

6app: Colorless crystals, mp 112–113°C (EtCN). IR (KBr) cm⁻¹: 3416 (OH of alcohol), 1573, 1503 (C=N), 1099 (C–N), 1253, 1099, 1032 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.17 (4H, m, H3′β, 5′β), 1.77 (6H, dm, J=12.8 Hz, H3′α, 5′α, 4′α), 1.82 (2H, br s, OH), 2.80 (4H, tm, J=13.1 Hz, H2′β, 6′β), 3.40 (3H, s, OCH₃), 3.49 (4H, d, J=6.1 Hz, H1), 3.71 (2H, t, J=5.2 Hz, H2‴), 4.42 (2H, t, J=5.2 Hz, H1‴), 4.76 (4H, br d, J=13.1 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.55 (C3′, 5′), 39.08 (C4′), 43.27 (C2′, 6′), 58.89 (OCH₃), 65.05 (C1‴), 67.54 (C1), 70.60 (C2‴), 165.80 (C2″, 4″), 170.73 (C6″). Positive-ion FAB-MS m/z: 382 (M+H⁺). HR-FAB-MS m/z: 382.2455 (Calcd for C₁₈H₃₂N₅O₄: 382.2454). Anal. Calcd for C₁₈H₃₁N₅O₄: C, 56.67; H, 8.19; N, 18.36. Found: C, 56.60; H, 8.17; N, 18.40.

6wpp: Colorless crystals, mp 262–264°C (EtOH). IR (KBr) cm⁻¹: 3425, 3286 (OH of alcohol), 2935 (NH), 1643, 1587, 1550 (C=O and C=N), 1263, 1043 (C–O of alcohol). ¹H-NMR (DMSO-d₆) δ: 1.05 (4H, m, H3′β, 5′β), 1.62 (2H, m, H4′), 1.67 (4H, br d, J=13.1 Hz, H3′α, 5′α,), 2.80 (4H, m, H2′β, 6′β), 3.25 (4H, dd, J=5.8, 5.5 Hz, H1), 4.43 (4H, dd, J=5.8, 5.5 Hz, H2′α, 6′α), 4.46 (2H, br s, OH), 10.27 (1H, br s, NH). ¹³C-NMR (DMSO-d₆) δ: 28.25 (C3′, 5′), 38.27 (C4′), 43.26 (C2′, 6′), 65.42 (C1), 157.44 (C4″, 6″), 159.82 (C2″). Positive-ion FAB-MS m/z: 324 (M+H⁺). HR-FAB-MS m/z: 324.2030 (Calcd for C₁₅H₂₆N₅O₃: 324.2036). Anal. Calcd for C₁₅H₂₅N₅O₃·0.3H₂O : C, 54.79; H, 7.85; N, 21.30. Found: C, 54.77; H, 7.76; N, 21.24.

7wap: mp 156–158°C. IR (KBr) cm⁻¹: 3480 (OH of alcohol), 2922 (NH), 1667, 1618, 1559, 1517, 1444 (C=O and C=N), 1295, 1131, 1095 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 0.88 (0.3H, br s, NH), 1.25 (2H, m, H3′β, 5′β), ca. 1.25 (0.7H, OH on C4″), 1.80 (3H, m, H3′α, 5′α, 4′), ca. 1.8 (1H, OH on C1), 2.90 (2H, t, J=13.1 Hz, H2′β, 6′β), 3.39 (3H, s, OCH₃), 3.51 (2H, d, J=6.1 Hz, H1), 3.69 (2H, t, J=4.9 Hz, H2‴), 4.50 (2H, t, J=4.9 Hz, H1‴), 4.77 (2H, br d, J=13.1 Hz, H2′α, 6′α), 4.88 (1H, br s, OH). ¹³C-NMR (CDCl₃) δ: 28.54 (C3′, 5′), 38.65 (C4′), 44.30 (C2′, 6′), 59.01 (OCH₃), 66.92 (C1‴), 67.08 (C1), 69.89 (C2‴), 159.68 (C4″), 162.35 (C2″), 164.10 (C6″). Positive-ion FAB-MS m/z: 285 (M+H⁺). HR-FAB-MS m/z: 285.1573 (Calcd for C₁₂H₂₁N₄O₄: 285.1563). Anal. Calcd for C₁₂H₂₀N₄O₄·0.25H₂O: C, 49.90; H, 7.15; N, 19.40. Found: C, 49.90; H, 7.32; N, 19.18.

To a solution of 1 (922 mg, 5.0 mmol) and DIPEA (2.58 g, 20.0 mmol) in dry THF (125 mL) was added aH (380 mg, 5.0 mmol) at 0°C under an N₂ atmosphere with stirring. After stirring for 15 min at 0°C and for 10 min at room temperature, additional DIPEA (2.58 g, 20.0 mmol) and aH (380 mg, 5.0 mmol) were added, and then the reaction mixture was stirred for 16 h at room temperature. (Step 2) To this yellow reaction mixture was added pH (1.15 g 10.0 mmol), and the resulting mixture was stirred at 70°C for 1 h under an N₂ atmosphere. After filtration of the separated amine hydrochloride (pH·HCl), the filtrate was evaporated and the residue was separated by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=95 : 4.7 : 0.3→93 : 6.6 : 0.4) to give 5rp (221 mg, 14%), 8ap (941 mg, 62%), 6app (253 mg, 13%), and 5pp⁷⁾ (22 mg, 1%). An analytical sample of 8ap was obtained by recrystallization from EtOH–H₂O.

8ap: Colorless crystals, mp 86–87°C (EtOH–H₂O). IR (KBr) cm⁻¹: 3416 (OH of alcohol), 1618, 1592 (C=N), 1090, 1055 (C–O of alcohol and ether), 794 (C–Cl). ¹H-NMR (CDCl₃) δ: 1.23 (2H, m, H3′β, 5′β), 1.57 (2H, br s, OH), 1.83 (3H, m, H3′α, 5′α, 4′α), 2.91 (2H, m, H2′β, 6′β), 3.41 (3H, s, OCH₃), 3.53 (2H, d, J=6.1 Hz, H1), 3.71 (2H, t, J=4.9 Hz, H2‴), 4.48 (2H, t, J=4.9 Hz, H1‴), 4.78 (2H, m, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.42 (C3′ or 5′), 28.50 (C5′ or 3′), 38.72 (C4′), 43.83 (C2′ or 6′), 43.97 (C6′ or 2′), 59.05 (OCH₃), 66.87 (C1‴), 67.22 (C1), 70.19 (C2‴), 165.11 (C2″), 170.54 (C4″), 170.89 (C6″). Positive-ion FAB-MS m/z: 303 (M+H⁺). HR-FAB-MS m/z: 303.1229 (Calcd for C₁₂H₂₀ClN₄O₃: 303.1224). Anal. Calcd for C₁₂H₁₉ Cl N₄O₃·0.1H₂O: C, 47.32; H, 6.35; N, 18.40. Found: C, 47.23; H, 6.22; N, 18.39.

5rp: ¹H-NMR (CDCl₃) δ: 1.95 (11H, m, CH₃×3, H3′β, 5′β), 1.62 (1H, br s, OH), 1.79 (3H, m, H4′α, 3′α, 5′α,), 2.82 (2H, m, H2′β, 6′β), 3.41 (2H, m, NCH₂CH₃), 3.52 (2H, d, J=6.1 Hz, H1), 4.74 (2H, m, H2′α, 6′α), 4.98 (1H, m, NCH<). ¹³C-NMR (CDCl₃) δ: 14.26 (NCH₂CH₃), 20.47 (NCH–CH₃), 28.55 (C3′, 5′), 36.61 (NCH₂CH₃), 39.01 (C4′), 43.42 (C2′, 6′), 46.06 (NCH–CH₃), 67.54 (C1), 164.05, 164.18 (C2″, 4″), 169.12 (C6″). Positive-ion FAB-MS m/z: 314 (M+H⁺). HR-FAB-MS m/z: 314.1748 (Calcd for C₁₄H₂₅ClN₅O: 314.1748).

These compounds were prepared by method A under the conditions shown in Table 1. Purification of the products by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=95 : 4.7 : 0.3) gave 7aap (60%), 6app (8%), and 6wpp (25%).

7aap: Pale yellow oil. IR (NaCl) cm⁻¹: 3420 (OH of alcohol), 1584, 1526 (C=N), 1118, 1036 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.19 (2H, m, H3′β, 5′β), 1.79 (3H, m, H3′α, 5′α, 4′α), 2.03 (1H, br s, OH), 2.86 (2H, tm, J=13.1 Hz, H2′β, 6′β), 3.40 (6H, s, OCH₃), 3.50 (2H, d, J=6.1 Hz, H1), 3.71 (4H, t, J=4.9 Hz, H2‴), 4.47 (4H, t, J=4.9 Hz, H1‴), 4.79 (2H, dm, J=13.1 Hz, H2′α, 6′α). ¹³C-NMR (CDCl₃) δ: 28.49 (C3′, 5′), 38.82 (C4′), 43.56 (C2′, 6′), 58.87 (OCH₃), 66.05 (C1‴), 67.19 (C1), 70.30 (C2‴), 166.26 (C2″), 171.72 (C4″, 6″). Positive-ion FAB-MS m/z: 343 (M+H⁺). HR-FAB-MS m/z: 343.1982 (Calcd for C₁₅H₂₇N₄O₅: 343.1981). Anal. Calcd for C₁₅H₂₆N₄O₅·0.3H₂O: C, 51.80; H, 7.71; N, 16.11. Found: C, 51.64; H, 7.85; N, 16.06.

These compounds were prepared from i-PrOH (bH) by method A under the conditions shown in Table 1. Purification of the products by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=95 : 4.7 : 0.3→85 : 14.5 : 0.5) gave 6bpp (84%) and 7wbp (5%) as a colorless solid and pale yellow solids, respectively. An analytical sample of 6bpp was obtained by recrystallization from EtCN as colorless crystals.

6bpp: mp 108–109°C (from EtCN). IR (KBr) cm⁻¹: 3387 (OH of alcohol), 1571, 1515 (C=N), 1253, 1112, 1036 (C–O). ¹H-NMR (CDCl₃) δ: 1.17 (4H, m, H3′β, 5′β), 1.33 (6H, d, J=6.1 Hz, CH₃), 1.72 (4H, m, H3′α, 5′α), 1.76 (2H, m, H4′α), 2.75 (2H, br s, OH), 2.78 (4H, t, J=12.5 Hz, H2′β, 6′β), 3.46 (4H, d, J=5.8 Hz, H1), 4.75 (4H, d, J=12.5 Hz, H2′α, 6′α), 5.23 (1H, qu, J=6.1 Hz, OCH<). ¹³C-NMR (CDCl₃) δ: 21.77 (CH₃), 28.44 (C3′, 5′), 38.87 (C4′), 43.11 (C2′, 6′), 67.17 (C1), 68.86 (OCH<), 165.66 (C2″, 4″), 170.15 (C6″). Positive-ion FAB-MS m/z: 366 (M+H⁺). HR-FAB-MS m/z: 366.2509 (Calcd for C₁₈H₃₂N₅O₃: 366.2505). Anal. Calcd for C₁₈H₃₁N₅O₃·0.25H₂O: C, 58.43; H, 8.58; N, 18.93. Found: C, 58.43; H, 8.81; N, 18.79.

7wbp: mp 203–205°C. ¹H-NMR (CDCl₃) δ: 1.2 (2H, m, H3′β, 5′β), 1.35 (6H, d, J=6.4 Hz, CH₃), 1.72 (2H, br s, OH), 1.85 (3H, m, H4′α, 3′α, 5′α), 2.89 (2H, m, H2′β, 6′β), 3.51 (2H, d, J=5.8 Hz, H1), 4.7, 4.9 (2H, br s, H2′α, 6′α), 5.29 (1H, qu, J=6.4 Hz, OCH<), 11.5 (1H, br s, NH). ¹³C-NMR (CDCl₃) δ: 21.63 (CH₃), 28.53 (C3′, 5′), 38.81 (C4′), 44.16 (C2′, 6′), 67.29 (C1), 72.25 (OCH<), 159.79, 161.51, 164.35 (C2″, 4″, 6″). ¹³C-NMR (DMSO-d₆) δ: 21.37 (CH₃), 28.41 (C3′, 5′), 38.26 (C4′), 43.46 (C2′, 6′), 65.38 (C1), 71.34 (OCH<), 156.83, 161.88, 163.39 (C2″, 4″, 6″). Positive-ion FAB-MS m/z: 269 (M+H⁺). HR-FAB-MS m/z: 269.1614 (Calcd for C₁₂H₂₁ N₄O₃: 269.1608).

This compound was prepared from bH by method A under the conditions shown in Table 1. Separation of the products by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=95 : 4.7 : 0.3→93 : 6.6 : 0.4) gave 7bbp (26%) and 6bpp (73%) as a pale yellow oil and pale yellow solid, respectively. An analytical sample of 7bbp was obtained by recrystallization from methylcyclohexane as pale yellow crystals.

7bbp: mp 95–96°C (from methylcyclohexane). IR (KBr) cm⁻¹: 3398 (OH of alcohol), 1580, 1540 (C=N), 1273, 1129 (C–O). ¹H-NMR (CDCl₃) δ: 1.20 (2H, m, H3′β, 5′β), 1.36 (12H, d, J=6.4 Hz, CH₃), 1.79 (4H, m, H3′α, 5′α, 4′α, OH), 2.85 (2H, dt, J=12.8, 2.4 Hz, H2′β, 6′β), 3.52 (2H, d, J=6.1 Hz, H1), 4.80 (2H, dm, J=13.4 Hz, H2′α, 6′α), 5.29 (1H, qu, J=6.4 Hz, OCH<). ¹³C-NMR (CDCl₃) δ: 21.90 (CH₃), 28.58 (C3′, 5′), 39.00 (C4′), 43.54 (C2′, 6′), 67.41 (C1), 70.08 (OCH<), 166.63 (C2″), 171.46 (C4″, 6″). Positive-ion FAB-MS m/z: 311 (M+H⁺). HR-FAB-MS m/z: 311.2080 (Calcd for C₁₅H₂₇N₄O₃: 311.2083). Anal. Calcd for C₁₅H₂₆N₄O₃·0.25H₂O: C, 58.04; H, 8.44; N, 18.05. Found: C, 57.84; H, 8.56; N, 18.02.

To a mixture of NaHCO₃ (2.52 g, 30 mmol) and dry bH (2.88 g, 48 mmol) in dry CHCl₃ (4 mL) was added compound 1 (1.84 g, 10 mmol) at room temperature with stirring. After stirring for 1 h at room temperature, the reaction mixture was refluxed for 17 h. After addition of CHCl₃ (10 mL), the resulting mixture was washed with 1% aqueous NaHCO₃ solution (10 mL×3) and the organic layer was dried with MgSO₄. After evaporation of the solvent, the residue was separated by centrifugal chromatography (n-hexane : EtOAc=95 : 5) to give 2b (583 mg, 28% yield) as a colorless oil.

2b: IR (NaCl) cm⁻¹: 2987 (CH), 1542, 1500 (C=N), 1100, 1036 (C–O of ether), 861, 807 (C–Cl). ¹H-NMR (CDCl₃) δ: 1.44 (6H, d, J=6.1 Hz, CH₃), 5.41 (1H, qu, J=6.1 Hz, O–CH<). ¹³C-NMR (CDCl₃) δ: 21.42 (CH₃), 74.90 (O–CH<), 170.51 (C2), 172.47 (C4, 6). The positive FAB mass spectra of this compound showed no significant molecular ion to determine its chemical formula, indicating the instability of the corresponding molecular ion in the mass range.

To a mixture of NaHCO₃ (5.04 g, 60 mmol) and dry bH (7.20 g, 120 mmol) in dry THF (40 mL) was added compound 1 (3.69 g, 20 mmol) at room temperature under an N₂ atmosphere with stirring. After stirring for 7 d at room temperature, the reaction mixture was filtrated over celite and the filtrate was evaporated. To the residual oil was added CH₂Cl₂ (50 mL) and the separated insoluble material was removed by filtration. The solvent was evaporated and the residual oil was separated by centrifugal chromatography (n-hexane : EtOAc=98 : 2→90 : 10) to give 3bb (79 mg, 2%), 4bbb (442 mg, 9%).

4bbb: Colorless crystals, mp 92–94°C. IR (KBr) cm⁻¹: 3433 (OH of H₂O), 1563 (C=N), 1148, 1095 (C–O of ether). ¹H-NMR (CDCl₃) δ: 1.38 (18H, d, J=6.4 Hz, CH₃), 5.35 (3H, qu, J=6.4 Hz, O–CH<). ¹³C-NMR (CDCl₃) δ: 21.81 (CH₃), 71.32 (O–CH<), 172.67 (C=N). Positive-ion FAB-MS m/z: 256 (M+H⁺). HR-FAB-MS m/z: 256.1658 (Calcd for C₁₂H₂₂N₃O₃: 256.1661). Anal. Calcd for C₁₂H₂₁N₃O₃·0.2H₂O: C, 55.67; H, 8.33; N, 16.23. Found: C, 55.73; H, 8.35; N, 16.37.

3bb: ¹H-NMR (CDCl₃) δ: 1.40 (12H, d, J=6.4 Hz, CH₃), 5.35 (2H, qu, J=6.4 Hz, O–CH<). ¹³C-NMR (CDCl₃) δ: 21.63 (CH₃), 71.97 (O–CH<), 171.63 (C2, 4), 172.61 (C6). Positive-ion FAB-MS m/z: 232 (M+H⁺). HR-FAB-MS m/z: 232.0852 (Calcd for C₉H₁₅ClN₃O₂: 232.0853).

This compound was prepared from 3-pentanol (cH) by method A under the conditions shown in Table 1. Purification of the product by flash chromatography (n-hexane : i-PrOH=90 : 10→65 : 35) gave 6cpp (1.42g, 72%) as a white solid. An analytical sample of 6cpp was obtained by recrystallization from EtCN as colorless crystals.

6cpp: mp 114–116°C (from EtCN). IR (KBr) cm⁻¹: 3399 (OH of alcohol), 1562, 1504 (C=N), 1249, 1112, 1035 (C–O). ¹H-NMR (CDCl₃) δ: 0.93 (6H, t, J=7.3 Hz, CH₃), 1.18 (4H, m, H3′β, 5′β), 1.59 (2H, br s, OH), 1.67 (4H, m, CH₂CH₃), 1.77 (6H, m, H3′α, 5′α, 4′α), 2.79 (4H, dt, J=13.1, 2.1 Hz, H2′β, 6′β), 3.50 (4H, d, J=5.8 Hz, H1), 4.77 (4H, d, J=13.1 Hz, H2′α, 6′α), 4.98 (1H, m, OCH<). ¹³C-NMR (CDCl₃) δ: 9.94 (CH₃), 26.57 (CH₂CH₃), 28.57 (C3′, 5′), 39.16 (C4′), 43.27 (C2′, 6′), 67.67 (C1), 78.40 (OCH<), 166.01 (C2″, 4″), 171.16 (C6″). Positive-ion FAB-MS m/z: 394 (M+H⁺). HR-FAB-MS m/z: 394.2820 (Calcd for C₂₀H₃₆N₅O₃: 394.2818). Anal. Calcd for C₂₀H₃₅N₅O₃·1.2H₂O: C, 57.86; H,9.08; N, 16.87. Found: C, 57.84; H, 8.87; N, 16.81.

To a solution of cH (882 mg, 10.0 mmol) in dry THF (30 mL) was added n-BuLi (1.6 mol/L n-hexane solution, 6.25 mL, 10.0 mmol) at room temperature under an an N₂ atmosphere. After stirring for 20 min at room temperature, a solution of compound 1 (922 mg, 5.0 mmol) in dry THF (10 mL) was added at room temperature. After additional stirring for 30 min at room temperature, the reaction mixture was refluxed for 2.5 h. After the solvent had been removed by evaporation, n-hexane (60 mL) and water (20 mL) were added. The organic layer was separated, dried (MgSO₄), and evaporated. (Step 2) The residual yellow oil was dissolved in dry CH₃CN (20 mL), and pH (922 mg, 8.0 mmol) was added, and the mixture was stirred for 30 min at room temperature. After filtration of the salt (pH·HCl) and evaporation of the solvent, the residue was separated by flash chromatography (n-hexane : i-PrOH : Et₂NH=130 : 20 : 0.1) to give 7ccp (1.33 g, 73%) as a colorless oil.

7ccp: ¹H-NMR (CDCl₃) δ: 0.94 (12H, t, J=7.5 Hz, H1‴, 5‴), 1.22 (2H, m, H3′β, 5′β), 1.54 (1H, br s, OH), 1.70 (8H, m, H2‴, 4‴), 1.80 (3H, m, 4′α, 3′α, 5′α), 2.85 (2H, tm, J=13.1 Hz, H2′β, 6′β), 3.52 (2H, d, J=5.8 Hz, H1), 4.80 (2H, dm, J=13.1 Hz, H2′α, 6′α), 5.04 (2H, qu, J=6.7 Hz, H3‴). ¹³C-NMR (CDCl₃) δ: 9.71 (C1‴, 5‴), 26.42 (C2‴, 4‴), 28.53 (C3′, 5′), 39.01 (C4′), 43.54 (C2′, 6′), 67.50 (C1), 79.29 (C3‴), 166.72 (C2″), 172.17 (C4″, 6″). Positive-ion FAB-MS m/z: 367 (M+H⁺). HR-FAB-MS m/z: 367.2710 (Calcd for C₁₉H₃₅N₄O₃: 367.2709).

This compound was prepared from cyclopentanol (dH) by method A under the conditions shown in Table 1. Purification of the product by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=97 : 2.7 : 0.3→90 : 9.5 : 0.5) gave 7ddp (25%) and 6dpp (67%) as a pale yellow oil and pale yellow solid, respectively. An analytical sample of 6dpp was obtained by recrystallization from CH₃CN as pale yellow crystals.

6dpp: mp 178–179°C (from CH₃CN). IR (KBr) cm⁻¹: 3427 (OH of alcohol), 1568, 1519 (C=N), 1253, 1105, 1039 (C–O). ¹H-NMR (CDCl₃) δ: 1.18 (4H, m, H3′β, 5′β), 1.56 (2H, m, H3‴, 4‴), 1.59 (2H, br s, OH), 1.71–1.87 (10H, m, H4′α, 3‴, 4‴, 3′α, 5′α, 2‴, 5‴), 1.94 (2H, m, H2‴, 5‴), 2.79 (4H, dt, J=12.8, 12.1 Hz, H2′β, 6′β), 3.50 (4H, d, J=5.8 Hz, H1), 4.78 (4H, br d, J=12.8 Hz, H2′α, 6′α), 5.33 (1H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 24.04 (C3‴, 4‴), 28.58 (C3′, 5′), 32.77 (C2‴, 5‴), 39.14 (C4′), 43.26 (C2′, 6′), 67.65 (C1), 78.65 (C1‴), 165.90 (C2″, 4″), 170.71 (C6″). Positive-ion FAB-MS m/z: 392 (M+H⁺). HR-FAB-MS m/z: 392.2662 (Calcd for C₂₀H₃₄N₅O₃: 392.2662). Anal. Calcd for C₂₀H₃₃N₅O₃: C, 61.36; H, 8.50; N, 17.89. Found: C, 61.34; H, 8.70; N, 17.89.

This compound was prepared from dH by method C under the conditions shown in Table 1. Purification of the product by flash chromatography (n-hexane : i-PrOH : Et₂NH=120 : 30 : 0.1) gave 7ddp (67%) as a colorless oil. An analytical sample of 7ddp was obtained by recrystallization from methylcyclohexane as colorless crystals. Formation of compound 5pp was also detected by TLC.

7ddp: mp 62–66°C (from methylcyclohexane). IR (KBr) cm⁻¹: 3333 (OH of alcohol), 1584, 1544 (C=N), 1273, 1130, 1108, 1046 (C–O). ¹H-NMR (CDCl₃) δ: 1.21 (2H, m, H3′β, 5′β), 1.58 (5H, m, OH, H3‴, 4‴), 1.79 (11H, m, H4′α, 3‴, 4‴, 3′α, 5′α, 2‴, 5‴), 1.95 (4H, m, H2‴, 5‴), 2.85 (2H, dt, J=13.2, 2.7 Hz, H2′β, 6′β), 3.51 (2H, d, J=6.1 Hz, H1), 4.80 (2H, d, J=15.6 Hz, H2′α, 6′α), 5.40 (2H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 23.98 (C3‴, 4‴), 28.56 (C3′, 5′), 32.75 (C2‴, 5‴), 39.00 (C4′), 43.55 (C2′, 6′), 67.49 (C1), 79.56 (C1‴), 166.52 (C2″), 171.64 (C4″, 6″). Positive-ion FAB-MS m/z: 363 (M+H⁺). HR-FAB-MS m/z: 363.2397 (Calcd for C₁₉H₃₁N₄O₃: 363.2396). Anal. Calcd for C₁₉H₃₀N₄O₃·0.4C₇H₁₄: C, 65.17; H, 8.93; N, 13.95. Found: C, 65.07; H, 9.10; N, 13.68.

This compound was prepared from cyclohexanol (eH) by method A under the conditions shown in Table 1. Purification of the product by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=930 : 66 : 4) gave 6epp (80%) as a yellow solid. Recrystallization from CH₃CN gave an analytically pure product 6epp.

6epp: mp 172–173°C (from CH₃CN). IR (KBr) cm⁻¹: 3466, 3275 (OH of alcohol) 1252, 1038 (C–O of ether). ¹H-NMR (CDCl₃) δ: 1.18 (4H, m, H3′β, 5′β), 1.25 (1H, m, H4‴), 1.35 (2H, m, H3‴, 5‴), 1.51 (2H, m, H2‴, 6‴), 1.58 (1H, m, H4‴), 1.7–1.9 (10H, m, H4′, 3′α, 5′α, 3‴, 5‴, OH), 2.03 (2H, m, H2‴, 6‴), 2.79 (4H, dt, J=12.8, 2.1 Hz, H2′β, 6′β), 3.50 (4H, d, J=5.8 Hz, H1), 4.76 (4H, d, J=13.4 Hz, H2′α, 6′ α), 4.91 (1H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 24.28 (C3‴, 5‴), 25.56 (C4‴), 28.58 (C3′, 5′), 31.87 (C2‴, 6‴), 39.11 (C4′), 43.24 (C2′, 6′), 67.60 (C1), 74.54 (C1‴), 165.95 (C2″, 4″), 170.38 (C6″). Positive-ion FAB-MS m/z: 406 (M+H⁺). HR-FAB-MS m/z: 406.2820 (Calcd for C₂₁H₃₆N₅O₃: 406.2818). Anal. Calcd for C₂₁H₃₅N₅O₃: C, 62.20; H, 8.70; N, 17.27. Found: C, 62.04; H, 8.86; N, 17.16.

These compounds were prepared from eH by method A under the conditions shown in Table 1. Purification of the product by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=950 : 47 : 30→850 : 145 : 5) gave 7eep (54%) as a yellow solid, 6epp (22%) as a yellow solid, and 7wep (2%) as a white solid. Recrystallization from EtCN gave an analytically pure product 7eep as pale yellow needles.

7eep: mp 155–156°C (from EtCN). IR (KBr) cm⁻¹: 3468 (OH of alcohol) 1149, 1107, 1040 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.20 (2H, m, H3′β, 5′β), 1.25 (2H, m, H4‴), 1.37 (4H, m, H3‴, 5‴), 1.54 (4H, m, H2‴, 6‴), 1.58 (2H, m, H4‴), 1.6 (1H, br s, OH), 1.80 (7H, m, H3′α, 4′, 5′α, 3‴, 5‴, OH), 2.05 (4H, m, H2‴, 6‴), 2.85 (2H, dt, J=13.1, 2.4 Hz, H2′β, 6′β), 3.52 (2H, d, J=5.8 Hz, H1), 4.79 (2H, dm, J=13.1 Hz, H2′α, 6′α), 4.97 (2H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 24.10 (C3‴, 5‴), 25.47 (C4‴), 28.58 (C3′, 5′), 31.79 (C2‴, 6‴), 38.99 (C4′), 45.53 (C2′, 6′), 67.50 (C1), 75.37 (C1‴), 166.69 (C2″), 171.48 (C4″, 6″). Positive-ion FAB-MS m/z: 391 (M+H⁺). HR-FAB-MS m/z: 391.2709 (Calcd for C₂₁H₃₅N₄O₃: 391.2709). Anal. Calcd for C₂₁H₃₄N₄O₃: C, 64.59; H, 8.78; N, 14.35. Found: C, 64.58; H, 8.80; N, 14.38.

7wep: ¹H-NMR (CDCl₃) δ: 1.15–1.3 (3H, m, 3′β, 5′β, 4‴), 1.3–1.4 (2H, m, H3‴, 5‴), 1.5–1.6 (3H, m, H2‴, 4‴, 6‴), 1.7–1.9 (5H, m, H3′α, 4′, 5′α, 3‴, 5‴), 1.96 (2H, m, H2‴, 6‴), 2.23 (2H, br s, OH), 2.88 (2H, br t, J=11.9 Hz, H2′β, 6′β), 3.51 (2H, d, J=5.8 Hz, H1), 4.65–4.95 (2H, br m, H2′α, 6′α), 5.19 (1H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 23.63 (C3‴, 5‴), 25.20 (C4‴), 28.55 (C3′, 5′), 31.28 (C2‴, 6‴), 38.72 (C4′), 44.15 (C2′, 6′), 67.09 (C1), 75.29 (C1‴), 159.98 (C2″), 161.75 (C4″ or 6″), 163.90 (C6″ or 4″). Positive-ion FAB-MS m/z: 309 (M+H⁺). HR-FAB-MS m/z: 309.1929 (Calcd for C₁₅H₂₅N₄O₃: 309.1927).

These compounds were prepared from cycloheptanol (fH) by method A under the conditions shown in Table 1. Purification of the products by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=950 : 47 : 3→930 : 66 : 4) gave 7ffp (38%) as a white solid, 6fpp (54%) as a white solid, and 7wfp (5%) as a reddish yellow solid. Recrystallization from EtCN gave an analytically pure product 6fpp as pale yellow crystals.

6fpp: mp 161–162°C (from EtCN). IR (KBr) cm⁻¹: 3455 (OH of alcohol) 1252, 1094, 1037 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.18 (4H, m, H3′β, 5′β), 1.45 (2H, m, H3‴, 6‴), 1.57 (4H, m, H4‴, 5‴), 1.66 (2H, br s, OH) 1.69–1.81 (10H, m, H3‴, 6‴, 4′, 3′α, 5′α, 2‴, 7‴), 2.05 (2H, m, H2‴, 7‴), 2.79 (4H, dt, J=13.1, 1.8 Hz, H2′β, 6′β), 3.51 (4H, d, J=5.8 Hz, H1), 4.77 (4H, dm, J=13.1 Hz, H2′α, 6′α), 5.09 (1H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 23.37 (C3‴, 6‴), 28.41 (C4‴, 5‴), 28.59 (C3′, 5′), 33.88 (C2‴, 7‴), 39.16 (C4′), 43.28 (C2′, 6′), 67.71 (C1), 77.05 (C1‴), 166.01 (C2″, 4″), 170.38 (C6″). Positive-ion FAB-MS m/z: 420 (M+H⁺). HR-FAB-MS m/z: 420.2974 (Calcd for C₂₂H₃₈N₅O₃: 420.2975). Anal. Calcd for C₂₂H₃₇N₅O₃: C, 62.98; H, 8.89; N, 16.69. Found: C, 62.91; H, 8.90; N, 16.74.

7wfp: Orange crystals, mp 186–188°C (from CH₃CN). IR (KBr) cm⁻¹: 3235 (OH of alcohol) 1297, 1103, 1045 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.23 (2H, m, 3′β, 5′β), 1.45 (2H, m, H3‴, 6‴), 1.57 (4H, m, H4‴, 5‴), 1.71 (2H, m, H3‴, 6‴), 1.7–1.9 (6H, m, OH, H2‴, 7‴, 4′, 3′α, 5′α), 2.01 (2H, m, H2‴, 7‴), 2.89 (2H, m, H2′β, 6′β), 3.52 (2H, m, H1), 4.72 and 4.92 (2H, br s, H2′α, 6′α), 5.18 (1H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 22.86 (C3‴, 6‴), 28.30 (C4‴, 5‴), 28.44 (C3′, 5′), 33.46 (C2‴, 7‴), 38.81 (C4′), 43.17 (C2′, 6′), 67.29 (C1), 79.83 (C1‴), 159.62 (C2″), 161.30 (C4″ or C6″), 164.43 (C6″ or C4″). Positive-ion FAB-MS m/z: 323 (M+H⁺). HR-FAB-MS m/z: 323.2087 (Calcd for C₁₆H₂₇N₄O₃: 323.2083). Anal. Calcd for C₁₆H₂₆N₄O₃: C, 59.61; H, 8.13; N, 17.38. Found: C, 59.44; H, 8.25; N, 17.47.

This compound was prepared from fH by method A under the conditions shown in Table 1. Purification of the product by flash chromatography (CH₂Cl₂ : 95% EtOH : 28% NH₃=950 : 47 : 3→930 : 66 : 4) gave 7ffp (52%) as a white solid and 6fpp (31%). Recrystallization from CH₃CN gave an analytically pure product 7ffp as colorless needles.

7ffp: mp 127–128°C (from CH₃CN). IR (KBr) cm⁻¹: 3363 (OH of ROH) 1252, 1185, 1123 (C–O of ROH and ether). ¹H-NMR (CDCl₃) δ: 1.20 (2H, m, H3′β, 5′β), 1.46 (4H, m, H3‴, 6‴), 1.58 (8H, m, H4‴, 5‴), 1.68–1.84 (12H, m, H3‴, 6‴, 2‴, 7‴, 4′, 3′α, 5′α, OH), 2.04 (4H, m, H2‴, 7‴), 2.84 (2H, dt, J=13.1, 1.8 Hz, H2′β, 6′β), 3.51 (2H, d, J=5.8 Hz, H1), 4.79 (2H, dm, J=13.1 Hz, H2′α, 6′α), 5.15 (2H, m, H1‴). ¹³C-NMR (CDCl₃) δ: 23.09 (C3‴, 6‴), 28.38 (C4‴, 5‴), 28.57 (C3′, 5′), 33.82 (C2‴, 7‴), 39.01 (C4′), 43.54 (C2′, 6′), 67.52 (C1), 77.84 (C1‴), 166.71 (C2″), 171.42 (C4″, 6″). Positive-ion FAB-MS m/z: 419 (M+H⁺). HR-FAB-MS m/z: 419.3018 (Calcd for C₂₂H₃₉N₅O₃: 419.3022). Anal. Calcd for C₂₃H₃₈N₄O₃: C, 66.00; H, 9.15; N, 13.39. Found: C, 65.79; H, 9.31; N, 13.35.

This compound was prepared from sesamol (gH) by method A under the conditions shown in Table 1. Purification of the product by centrifugal chromatography (CH₂Cl₂ : EtOH=97 : 3) gave 7ggp (52%) as a white solid and 6gpp (16%) as a white solid. Recrystallization from EtCN gave an analytically pure product 7ggp as colorless crystals.

7ggp: mp 201–203°C (from EtCN). IR (KBr) cm⁻¹: 3377 (OH of alcohol), 1242, 1177, 1137, 1038 (C–O of alcohol and ether). ¹H-NMR (CDCl₃) δ: 1.68 (2H, m, H3′β, 5′β), 1.34 (1H, br s, OH), 1.75 (3H, m, H4′α, 3′α, 5′α), 2.82 (2H, dt, J=12.8, 2.1 Hz, H2′β, 6′β), 3.51 (2H, d, J=5.8 Hz, H1), 4.62 (2H, d, J=13.4 Hz, H2′α, 6′α), 5.97 (4H, s, H2‴), 6.59 (2H, dd, J=8.2, 2.1 Hz, H6‴), 6.66 (2H, d, J=2.1 Hz, H4‴), 6.74 (2H, d, J=8.2 Hz, H7‴). ¹³C-NMR (CDCl₃) δ: 28.45 (C3′, 5′), 38.81 (C4′), 43.75 (C2′, 6′), 67.35 (C1), 101.58 (C2‴), 104.04 (C4‴), 107.68 (C7‴), 114.11 (C6‴), 144.98 (C7‴a), 146.61 (C5‴), 147.73 (C3‴a), 166.53 (C2″), 172.53 (C4″, 6″). Positive-ion FAB-MS m/z: 467 (M+H⁺). HR-FAB-MS m/z: 467.1512 (Calcd for C₂₃H₂₃N₄O₇: 467.1567). Anal. Calcd for C₂₃H₂₂N₄O₇: C, 59.22; H, 4.75; N, 12.01. Found: C, 59.14; H, 4.82; N, 12.02.

The antiviral activities of synthesized compounds were measured by using a plaque reduction assay¹⁶⁾ as described in our previous paper.¹⁹⁾ Results for antiviral activity (EC₅₀) and cytotoxicity (IC₅₀) with Vero cells are summarized in Table 2.

• 1) Balzarini J., Antiviral Res., 71, 237–247 (2006), and related references cited therein.
• 2) Li L., Jose J., Xiang Y., Kuhn R. J., Rossmann M. G., Nature (London), 468, 705–708 (2010).
• 3) Trouche N., Wieckowski S., Sun W., Chaloin O., Hoebeke J., Fournel S., Guichard G., J. Am. Chem. Soc., 129, 13480–13492 (2007).
• 4) Contreras J.-M., Sippl W., “The Practice of Medicinal Chemistry,ˮ 3rd ed., ed. by Wermuth C. G., Elsevier Academic Press, London, 2008, pp. 380–414. ISBN: 978-0-12-374194-3
• 5) Gibson S. E., Castaldi M. P., Angew. Chem. Int. Ed., 45, 4718–4720 (2006).
• 6) Humblet V., Misra P., Bhushan K. R., Nasr K., Ko Y.-S., Tsukamoto T., Pannier N., Frangioni J. V., Maison W., J. Med. Chem., 52, 544–550 (2009), and related references cited therein.
• 7) Mibu N., Yokomizo K., Takemura S., Ueki N., Itohara S., Zhou J., Miyata T., Sumoto K., Chem. Pharm. Bull., 61, 823–833 (2013).
• 8) Mibu N., Aki H., Ikeda H., Saito A., Uchida W., Yokomizo K., Zhou J., Miyata T., Sumoto K., J Therm Anal Calorim, 113, 1015–1018 (2013).
• 9) Fujisaki F., Aki H., Naito A., Fukami E., Kashige N., Miake F., Sumoto K., Chem. Pharm. Bull., 62, 429–438 (2014), and related references cited therein.
• 10) Jastrzabek K. G., Subiros-Funosas R., Albericio F., Kolesinska B., Kaminski Z. J., J. Org. Chem., 76, 4506–4513 (2011).
• 11) Kamiński Z. J., Markowicz S. W., Kolesińska B., Martynowski D., Główka M. L., Synth. Commun., 28, 2689–2696 (1998).
• 12) Darcos V., Griffith K., Sallenave X., Desvergne J.-P., Guyard-Duhayon C., Hasenknopf B., Bassani D. M., Photochem. Photobiol. Sci., 2, 1152–1161 (2003).
• 13) Spielman M. A., Close W. J., Wilk I. J., J. Am. Chem. Soc., 73, 1775–1777 (1951). Compound 4bbb was reported as mp 102–103°C.
• 14) Hedayatullah M., Lion C., Ben Slimane A., Da Conceiqao L., Nachawati I., Heterocycles, 51, 1891–1896 (1999).
• 15) After chromatography of the reaction products, purification of compound 7ddp by recrystallization was employed, but purification of the product 7ccp was very difficult and gave an oily material containing a few contaminants that had similar chemical properties. A small amount of compound Cc or Cd shown below was detected as a contaminant in the obtained product 7ccp or 7ddp. We are considering investigation of further improvement in the purification stages of the products. The results of biological evaluations and the calculated log P value for compound 6cpp are shown in Table 2.
• 16) Schinazi R. F., Peters J., Williams C. C., Chance D., Nahmias A. J., Antimicrob. Agents Chemother., 22, 499–507 (1982).
• 17) Ikeda T., Yokomizo K., Okawa M., Tsuchihashi R., Kinjo J., Nohara T., Uyeda M., Biol. Pharm. Bull., 28, 1779–1781 (2005).
• 18) Log P was calculated by using ChemBioDraw 13.0.2.3021.
• 19) Mibu N., Yokomizo K., Kashige N., Miake F., Miyata T., Uyeda M., Sumoto K., Chem. Pharm. Bull., 55, 111–114 (2007).